System and method for spatially profiling a distribution of hydrophobicity of a transmembrane protein

ABSTRACT

A system and method for spatially profiling a distribution of hydrophobicity of a transmembrane protein includes a scaler which generates scaled hydrophobicity values for the transmembrane protein, and a profiler which spatially profiles a hydrophobicity distribution for the transmembrane protein based on the scaled hydrophobicity values.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates to a system and method for spatiallyprofiling a hydrophobicity distribution for a transmembrane protein, andmore particularly, to a system for spatially profiling a hydrophobicitydistribution for a transmembrane protein based on scaled hydrophobicityvalues for the transmembrane protein.

2. Description of the Related Art

The distribution of residue hydrophobicity from protein interior toexterior has been a subject of continued interest. The identification ofthe hydrophobic core of soluble globular proteins has provided a keyfeature relating an amino acid attribute to tertiary protein structure.Furthermore, a detailed relationship between the hydrophobic characterof a local sequence of amino acids and variations of its proximity tothe protein exterior has been described. The spatial segregation ofresidues, dependent upon amino acid hydrophobicity, has also assistedwith validating predicted native protein structures, as well as withidentifying the origin of nucleation sites during the initiation ofprotein folding.

While there has been a general consensus of opinion concerning theoverall hydrophobic spatial distribution of the residues of solubleproteins, opinion concerning the hydrophobic distribution of theresidues of transmembrane protein structures has had a more variedhistory. Early work had suggested that Bacteriorhodopsin was an“inside-out” protein. The terminology, “inside-out”, referred to areversed sense of the hydrophobic distribution within the lipid bilayerfrom that of soluble globular proteins, namely, that the interior wascomposed of hydrophilic residues and the exterior of hydrophobicresidues. Apparently the “inside-out” model of membrane proteinstructure is no longer accepted.

Notwithstanding, a relatively recent calculation that utilizessolvent-lipid accessibility, as have prior investigations, and purportsto discredit the “inside-out” hypothesis, actually attempts to discredita more general hypothesis, namely, that residues of greater hydrophobiccharacter have a statistical preference to reside nearer the proteinlipid interface. This latter hypothesis is independent of the residuecharacter of the protein interior, being a statement of variations aboutthe mean or average residue hydrophobicity of the distribution, whateverthat distribution might be.

However, conventional systems and methods do not attempt to profile thedistribution of hydrophobicity of transmembrane proteins. Thus,conventional systems and methods do not enable a determination of thespatial distribution of hydrophobicity within the interior of astructure, where the residue solvent-lipid exposure either vanishes oris minimal.

SUMMARY OF THE INVENTION

In view of the foregoing and other problems, disadvantages, anddrawbacks of the aforementioned assemblies and methods, it is a purposeof the exemplary aspects of the present invention to provide a systemand method for spatially profiling a hydrophobicity distribution for atransmembrane protein.

A hallmark of soluble globular protein tertiary structure is ahydrophobic core and a protein exterior populated predominantly byhydrophilic residues. Recent hydrophobic moment profiling of the spatialdistribution of thirty globular proteins of diverse size and structurehad revealed features of this distribution that were comparable.Analogous profiling of the hydrophobicity distribution of the α-helicalburied bundles of several transmembrane proteins, as the lipid/proteininterface is approached from within the bilayer, reveals spatialhydrophobicity profiles that contrast with those obtained for thesoluble proteins. The calculations enabling relative changes ofhydrophobicity to be simply identified over the entire spatial extent ofthe multimer within the lipid bilayer, show the accumulated zero-ordermoments of the bundles to be mainly inverted with respect to that foundfor the soluble proteins. This indicates a statistical increase in theaverage residue hydrophobic content as the lipid bilayer is approached.

This result differs from that of a relatively recent calculation andqualitatively agrees with earlier calculations involving lipid exposedand buried residues of the α-helices of transmembrane proteins. Spatialprofiling, over the entire spatial extent of the multimer with scaledvalues of residue hydrophobicity, further provides information that isnot available from calculations utilizing lipid exposure alone.

An exemplary aspect of the present invention includes a system forspatially profiling a hydrophobicity distribution for a transmembraneprotein (e.g., a transmembrane protein formed in a lipid bilayer). Thesystem includes a scaler which generates scaled hydrophobicity valuesfor the transmembrane protein, and a profiler which spatially profiles ahydrophobicity distribution for the transmembrane protein based on thescaled hydrophobicity values. The system may also include a databasewhich stores data pertaining to a structure of the transmembraneprotein, and an input device for inputting and manipulating the data,and a display device (e.g., user interface) for viewing and manipulatingdata in the system.

Further, the scaled hydrophobicity values may include shifted and scaledhydrophobicity values of residue hydrophobicity. More specifically, thescaled hydrophobicity values may be derived from data pertaining to astructure of the transmembrane protein.

In addition, the profiler may spatially profile the hydrophobicitydistribution over a spatial extent of the transmembrane protein.Further, the profiler may include an identifier which identifies aresidue external to the membrane, using at least one of hydrophobicitysliding window analysis and visual inspection. The profiler may furtherinclude a residue remover which removes the residue to obtain atruncated structure, and a calculator which calculates a residuecentroid of each residue side-chain of the truncated structure to obtaina geometric center of a distribution of the residue centroids. Inaddition, the calculator may calculate a hydrophobicity profile about anaxis through the geometric center and normal to a plane of the membrane,using a predetermined profiling geometry. Further, the profiler andscaler, and any components of the features may be integrally formed asone or more processors operatively coupled to form the presentinvention.

For example, the predetermined profiling geometry may be selected toapproximate an overall external shape of the transmembrane proteinwithin a lipid bilayer, and generate a series of nested shapesconsistent with the profiling geometry. Further, the series of nestedshapes may provide contours about the axis that correlate with a lipidexposure for the residues proximate to a protein/lipid boundary. Forexample, the structure of the transmembrane protein may be asymmetricalabout the axis, and the predetermined profiling geometry may include anelliptical cylinder.

Further, the scaler may scale the hydrophobicity values to obtain thescaled hydrophobicity values, using a scale that similarly segregatesamino acid values of hydrophobicity into a polar, polar uncharged andpolar charged residues. The scaler may also shift the scaledhydrophobicity values to provide shifted and scaled values having avalue of zero hydrophobicity when a predetermined number of residues ofeach truncated structure are collected, the shifted and scaled valuesbeing scaled to provide a standard deviation of unity for each truncatedstructure.

Further, the profiler may accumulate the shifted and scaled values ofresidue hydrophobicity as a function of an increasing size of eachnested shape of a profiling geometry until a largest shape encapsulatesa predetermined number of the residues, to generate an accumulatedspatial distribution of residue hydrophobicity given by a function H(d)which is a sum of residue hydophobicity values within the circular,elliptical or conical cylinder of radius d. The profiler may, therefore,obtain a hydrophobicity profile by calculating the values of H(d).

For example, if H(d) increases, the average hydrophobic value of theresidues collected over a shell of width one or more Angstroms isgreater than the average value of residue hydrophobicity for the entirestructure. However, if H(d) decreases, the average hydrophobic value ofthe residues collected over a shell of width one or more Angstroms isless than the average value of residue hydrophobicity for the entirestructure.

Further, the accumulated spatial distribution may include a set ofsequential values of accumulated residue hydrophobicity with increasingdistance from a center of the structure to the protein/lipid interfacewithin a bilayer, the set of sequential values comprising a zero-ordermoment profile of the residue hydrophobicity from the interior to theexterior of the structure.

Another exemplary aspect of the present invention includes a method ofspatially profiling a hydrophobicity distribution for a transmembraneprotein. The method includes scaling hydrophobicity values for thetransmembrane protein to generate scaled hydrophobicity values, andspatially profiling a hydrophobicity distribution for the transmembraneprotein based on the scaled hydrophobicity values.

Specifically, the spatially profiling may include identifying a residueexternal to the membrane, using hydrophobicity sliding window analysisand by visual inspection, removing the residue to obtain a truncatedstructure, calculating a residue centroid of each residue side-chain ofthe truncated structure to obtain a geometric center of a distributionof the residue centroid. and/or calculating a hydrophobicity profileabout an axis through the geometric center and normal to the plane ofthe membrane, using a predetermined profiling geometry.

Further, scaling the hydrophobicity values may include scalinghydrophobicity values to obtain the scaled hydrophobicity values, usinga scale that similarly segregates amino acid values of hydrophobicityinto a polar, polar uncharged and polar charged residues. In addition,scaling the hydrophobicity values may further include shifting thescaled hydrophobicity values to provide shifted and scaled values havinga value of zero hydrophobicity when a predetermined number of residuesof each truncated structure are collected, the shifted and scaled valuesbeing scaled to provide a standard deviation of unity for each truncatedstructure.

Further, spatially profiling may further include accumulating theshifted and scaled values of residue hydrophobicity as a function ofincreasing size of each nested shape of the profiling geometry until alargest shape encapsulates a predetermined number of the residues, togenerate an accumulated spatial distribution of residue hydrophobicitygiven by the function H(d) which is the sum of the values of residuehydophobicity within the circular, elliptical or conical cylinder ofradius d. Spatially profiling may also include obtaining ahydrophobicity profile by calculating the values of H(d).

Another exemplary aspect of the present invention includes aprogrammable storage medium tangibly embodying a program ofmachine-readable instructions executable by a digital processingapparatus to perform the inventive method of spatially profiling ahydrophobicity distribution for a transmembrane protein.

Another exemplary aspect of the present invention includes a method ofdeploying computing infrastructure in which computer-readable code isintegrated into a computing system, such that the code and the computingsystem combine to perform the inventive method of spatially profiling ahydrophobicity distribution for a transmembrane protein.

With its unique and novel features, the present invention provides asystem and method which enables a determination of the spatialdistribution of hydrophobicity over an entire multimeric extent, notonly in the region proximate to the protein lipid interface.

BRIEF DESCRIPTION OF THE DRAWINGS

The foregoing and other exemplary aspects and advantages will be betterunderstood from the following detailed description of the exemplaryembodiments of the invention with reference to the drawings, in which:

FIG. 1 illustrates a system 100 for spatially profiling a hydrophobicitydistribution for a transmembrane protein, according to an exemplaryembodiment of the present invention;

FIG. 2 illustrates the truncated structure of the seven A-chain helicesof Bacteriorhodopsin, 1 C3W;

FIGS. 3A-3D illustrate a distribution of residue centroids andhydophobicity profiles of Bacteriorhodopsin, 1C3W, according to anexemplary embodiment of the present invention;

FIGS. 4A-4D illustrate a distribution of residue centroids andhydophobicity profiles of the subunit C of the ATP synthase fromEscherichia coli, 1 C 17, according to an exemplary embodiment of thepresent invention;

FIG. 5 illustrates a view, along the normal to the membrane surface, ofthe helices of the truncated multimer of the mechanosensitive ionchannel, 1 MSL;

FIGS. 6A-6D illustrate a distribution of residue centroids andhydophobicity profiles of the gated mechanosensitive ion channel fromMycobacterium tuberculosis, 1MSL, according to an exemplary embodimentof the present invention;

FIGS. 7A-7D illustrate a distribution of residue centroids andhydophobicity profiles of the potassium ion channel from Streptomyceslividans, 1K4D, according to an exemplary embodiment of the presentinvention;

FIG. 8A-8D illustrate a distribution of residue centroids andhydophobicity profiles of the photosynthetic reaction center,Rhodobacter sphaeroides, 1PCR, according to an exemplary embodiment ofthe present invention;

FIGS. 9A-9D illustrate a distribution of residue centroids andhydophobicity profiles of the Cytochrome-C Oxidase, 1EHK, according toan exemplary embodiment of the present invention;

FIGS. 10A-10D illustrate a distribution of residue centroids andhydophobicity profiles of the Aqpl Water Channel, 1J4N, according to anexemplary embodiment of the present invention;

FIGS. 11A-11D illustrate a distribution of residue centroids andhydophobicity profiles of the Bacterial Abc Transporter, 1L7V, accordingto an exemplary embodiment of the present invention;

FIGS. 12A-12D illustrate a distribution of residue centroids andhydophobicity profiles of E. Coli Quinol-Fumarate Reductase, 1KF6,according to an exemplary embodiment of the present invention;

FIGS. 13A-13D illustrates a distribution of residue centroids andhydophobicity profiles of the Cytochrome Bc 1 Complex, 1 BE3, accordingto an exemplary embodiment of the present invention;

FIGS. 14A-14D illustrate a distribution of residue centroids andhydophobicity profiles of the Photosynthetic Reaction Center:Photosystem I, 1JB0, according to an exemplary embodiment of the presentinvention;

FIGS. 15A-15F illustrate the hydrophobicity density, ρ(d), as a functionof distance, d, from the protein interior of the four transmembranebundles of, 1 C3W, 1C17, 1 MSL, 1 K4d and of the soluble proteins, 1AKZand 3PBG, according to an exemplary embodiment of the present invention;

FIG. 16 provides Table 1 which lists the shifted and normalized valuesof amino acid hydrophobicity that had provided the values of thedensities, ρ(d);

FIG. 17 is a flowchart illustrating a method of spatially profiling ahydrophobicity distribution for a transmembrane protein, according to anexemplary embodiment of the present invention;

FIG. 18 illustrates a typical hardware configuration which may be usedto implement the inventive system and method for spatially profiling ahydrophobicity distribution for a transmembrane protein, according to anexemplary embodiment of the present invention; and

FIG. 19 illustrates a programmable storage medium which may be used toperform a method for spatially profiling a hydrophobicity distributionfor a transmembrane protein, according to an exemplary embodiment of thepresent invention.

DETAILED DESCRIPTION OF THE EMBODIMENTS OF THE INVENTION

Referring now to the drawings, FIG. 1 illustrates a system 100 forspatially profiling the distribution of hydrophobicity of transmembraneproteins. As shown in FIG. 1, the system 100 includes a scaler 110 whichgenerates scaled hydrophobicity values for the transmembrane protein,and a profiler 115 which spatially profiles a hydrophobicitydistribution for the transmembrane protein based on the scaledhydrophobicity values (e.g., shifted and scaled hydrophobicity values)the transmembrane protein.

The system 100 may also include an input device 120 for inputting data(e.g., structural data) pertaining to the transmembrane protein. Thesystem 100 may also include a memory device 130 (e.g., database) forstoring data used by the system 100, and data generated by the system100. The system 100 may also include a display device 140 which may beused to display the results of the calculations performed by theprocessor 110, and a printing device 150 which may be used to printthose results.

Motivated by the problems of conventional systems and methods, theinventor investigated the distribution of residue hydrophobicity ofseveral transmembrane proteins from a point of view which is differentfrom conventional methods. The direct spatial profiling of the residuedistribution of the multimeric protein structures was performed withoutreference to residue solvent-lipid accessibility. Profiling in thismanner also enables a determination of the spatial distribution ofhydrophobicity within the interior of the structure, where the residuesolvent-lipid exposure either vanishes or is minimal.

A recent calculation examined the spatial distribution of the residuehydrophobicity of soluble globular proteins. It was shown that shiftingthe scale of residue hydrophobicity, such that the total residuehydrophobicity of each structure vanished, enabled variations of thespatial distribution of residue hydrophobicity about the mean of thedistribution to be simply identified. It also enabled a comparison to bemade between the hydrophobicity distributions of different proteins overtheir different length scales.

Such procedure is, therefore, appropriate for examining spatialvariations of the hydrophobic content of the residues of transmembraneproteins. It not only addresses the question of the hydrophobicstatistical preference of residues directly, but also providesinformation over the entire multimeric extent, not only in the regionproximate to the protein lipid interface.

The hydrophobicity profiles of globular proteins had revealed twospatial regions delineating the hydrophobic core and hydrophilicexterior. The profiles of the multimers comprised of transmembraneα-helices, while mainly inverted with respect to the profiles of theglobular proteins, do not always exhibit such uniform delineation. Thespatial profiling of structures about the normal to the plane of thelipid bilayer, yields features related to structural details within theinterior of the helical bundle as well as features related to exteriorlocal structural details that are not characteristic of the entireprotein-lipid periphery.

On the other hand, the major fraction of residues that are proximate tothe protein lipid interface for all of the α-helical structuresinvestigated were shown to exhibit a statistical increase in residuehydrophobic content as the interface is approached. The results,therefore, qualitatively agree with previous calculations involvingsurface exposed and buried residues. This increase in residuehydrophobic content, as the protein-lipid interface is approached is,however, more modest than the converse variation observed for thesoluble globular proteins.

The inventor conducted experiments in which eleven transmembrane proteinstructures with a SCOP ‘membrane all- α fold’ classification weredownloaded from the Protein Data Bank. The light-driven ion pump,Bacteriorhodopsin, 1C3W, and the photosynthetic reaction center,Rhodobacter sphaeroides, 1PCR, were chosen since they had been thesubject of previous discussion focused on their spatial distribution ofa polar and polar residues.

Three of the structures were chosen due to their symmetric, as well asdiverse multimeric geometries. The gated mechanosensitive ion channelfrom Mycobacterium tuberculosis, 1MSL, was chosen due to itsinterestingly entwined multimeric cylindrical symmetry with helices thatare canted significantly with respect to the surface of the lipidbilayer. The subunit C of the ATP synthase from Escherichia coli, 1 C17, was chosen due to its cylindrically symmetric set of helicesapproximately perpendicular to the lipid bilayer. The potassium ionchannel from Streptomyces lividans, 1K4D, was chosen due to the overallconical geometry of its membrane-spanning segment.

Six structures lacking symmetry about an axis normal to the plane of thebilayer were additionally chosen. The six are the Cytochrome-C Oxidase,1EHK, the Aqpl Water Channel, 1J4N, the Bacterial Abc Transporter, 1L7V,E. Coli Quinol-Fumarate Reductase, 1KF6, the Cytochrome Bcl Complex,1BE3, and the Photosynthetic Reaction Center: Photosystem I, 1JB0.

Prior to calculation, residues presumed to be external to the membranewere removed. These residues were identified by hydrophobicity slidingwindow analysis and by visual inspection. The final truncated proteinstructures were composed of a majority of α-helices with five turns orgreater.

FIG. 2 illustrates the truncated structure of the seven helices ofBacteriorhodopsin after residue elimination. The largest structure, thephotosynthetic reaction center, 1JB0, contained thirty-one truncatedhelices.

The centroid of each residue side-chain of the truncated structure iscalculated and the geometric center of the distribution of residuecentroids obtained. The shape of a profiling geometry (e.g., sphere,cylinder, cone, ellipsoid, etc.) is then chosen and the hydrophobicityprofile is calculated about the axis through the geometric center andnormal to the plane of the membrane.

For the structures exhibiting symmetry about the profiling axis, namely1 C3W, 1 C 17, 1 MSL, and 1 MD, the choice of the profiling geometry waschosen to approximate the overall external shape of the multimer withinthe lipid bilayer and a series of nested shapes consistent with thisgeometry were generated. This choice of nested shapes provides contoursabout the profiling axis that correlate with the lipid exposure forresidues proximate to the protein lipid boundary.

For the structures with no symmetry about the profiling axis, anelliptical cylinder was chosen for profiling. At the greatest distancesfrom the profiling axis, averages with ellipses over an irregulardistribution of protein centroids will have contributions from localregions that are most distant from the profiling axis. This contrastswith the averages over the symmetric structures where at large distancesfrom the profiling axis, symmetrically related features that are notspatially contiguous are included in the average. Certain consequencesof averaging over the structures lacking symmetry are discussed below.

Results are obtained for the Eisenberg and for the GES scales of residuehydrophobicity. The GES scale was chosen since it had been used in theprevious calculation alluded to. While these two particular scales havebeen used, the overall qualitative features described below would berelatively insensitive to the choice of any scale that would similarlysegregate the amino acid values of hydrophobicity into a polar, polaruncharged and polar charged residues.

The scales are shifted to provide a value of zero hydrophobicity whenall residues of each truncated structure are collected. The shiftedvalues are also scaled to provide a standard deviation of unity for eachstructure. Since the average value of residue hydrophobicity of theentire structure is then zero, this shift of the scale enables a simpleinterpretation of the changes in the accumulated values of residuehydrophobicity with increasing distance from the protein interior.

If the value increases with increasing distance from the interior,residues of greater hydrophobic content than the average of the entirestructure have been collected. If the value decreases with increasingdistance from the interior, residues of lesser hydrophobic content havebeen collected. Shifting the scale of residue hydrophobicity in thismanner provides a baseline for comparison of changes in the spatialdistribution of residue hydrophobicity of the truncated structures. Itenables concise quantitative statements of the spatial changes inresidue hydrophobicity that are independent of the overall hydrophobiccontent of the structures as well as enabling a comparison of thesechanges over the spatial extent of different structures.

With the choice of a profiling geometry the values of residuehydrophobicity are then accumulated as a function of increasing size ofeach nested shape of the profiling geometry until the largest shapeencapsulates all of the residues. The accumulated spatial distributionof residue hydrophobicity, or accumulated zero-order moment profile, isgiven by the function H(d). H(d) is the sum of the values of residuehydophobicity within the circular, elliptical or conical cylinder ofradius d.

${H(d)} = \begin{matrix}\; & h_{i}^{\prime} \\i & d\end{matrix}$

The h′_(i) are the shifted and scaled values of hydrophobicity of theith residue.

The hydrophobicity profile is obtained by calculating the values of H(d)in steps. For example, the steps may be in one or more Angstroms. Aspreviously noted, the changes in H(d) are interpreted simply for eachincreasing value of d. If H(d) increases, the average hydrophobic valueof the residues collected over the shell (e.g., of width one or moreAngstroms) is greater than the average value of residue hydrophobicityfor the entire structure. If H(d) decreases, residues of lesser thanaverage hydrophobic value have been collected. Any subsequent commentmade with regard to increasing or decreasing hydrophobic residue contentmay then be made with respect to the average value of residuehydrophobicity of the entire structure.

Collecting the values of residue hydrophobicity in this manner providesa set of sequential values of accumulated residue hydrophobicity withincreasing distance from the center of the structure to theprotein/lipid interface within the bilayer. These values may form azero-order moment profile of the residue hydrophobicity from theinterior to the exterior of the structure.

Such a profile has been previously obtained for thirty soluble globularproteins. However, a second-order moment, which had been used to amplifythe distance dependence of the hydrophobicity distribution, and hadprovided the quasi-invariant hydrophobic-ratio of distances for solubleglobular proteins, is not necessarily utilized in the present invention.

FIGS. 3A-D, 4A-D, and 6A-D through 14A-D graphically illustrate thehydrophobicity profiles, H(d), for the structures and the distributionof residue centroids with elliptical or circular boundaries that delimitregions of contrasting behavior. The shapes are mainly inverted withrespect to the shapes obtained for the thirty soluble globular proteinspreviously investigated.

The interior regions exhibit diverse behavior, either populated, onaverage, by residues of greater than, roughly equal to or of lesserhydrophobic content than the average of the entire structure. All of theprofiles display an intermediate spatial region of decreasinghydrophobic content. At the most distant values from the interior, themajority of the profiles increase, on average, with increasing distance.The few profiles that do not, namely, profiles of the structures 1KF6, 1BE3, and 1JB0, reflect averages over local regions of the interface andnot averages over a major portion of the interface. The average over themajor fraction of the interface near the protein-lipid boundary shows astatistical increase in residue hydrophobic content with decreasingdistance to the protein-lipid interface for all eleven structures.

FIGS. 3A-3D illustrate a distribution of residue centroids andhydophobicity profiles of Bacteriorhodopsin, 1C3W. FIG. 3A illustrates aview of along the C3 symmetry axis normal to the plane of the membrane.FIG. 3B illustrates a view canted by 80 degrees from the symmetry axis.FIG. 3C illustrates the hydrophobicity profile, H(d), calculated withthe Eisenberg hydrophobicity scale. FIG. 3D illustrates thehydrophobicity profile, H(d), calculated with the GES hydrophobicityscale. The solid and dashed lines in FIGS. 3C and 3D are calculated insteps of one and two Angstroms, respectively;

Profile features will be related to the structural features of theα-helical bundles for the structures exhibiting symmetry.Bacteriorhodopsin (1 C3W) has been profiled with a series of nestedcylinders of varying radii. FIG. 3A is a view along the C3 symmetry axisthat lies close to the perpendicular to the plane of the membrane. Thedistribution of residue centroids is shown along with three cylindricalcircular crosssections of different radii. The largest cylinder with aradius of 29 Angstroms encloses all residue centroids. FIG. 3B is a viewthat is canted by 80 degrees from this axis. Three circular crosssections of the largest cylinder are shown in FIG. 3B.

FIGS. 3C and 3D show H(d) calculated with the Eisenberg and the GESresidue hydrophobicity scales, respectively. The general trends withincreasing radial distance from the cylindrical axis are similar. Thefirst six hydrophobic residues are collected at eight Angstroms. Thereis a subsequent increase in H(d), indicating the collection of residuesof increasing hydrophobic content within the protein interior.

In the range of 15 to 20 Angstroms, the accumulation of hydrophilicresidues and diminishing accumulation of hydrophobic residues isresponsible for the plunge to negative values. This range of distancesspans the range between the two inner circles of FIG. 3A, whichdelineate the region between the inner and outer nested bundles ofα-helices. In this region are the water bound molecules, the retinalSchiff bases, and three arginine, lysine, and aspartic acid residues.From 24 Angstroms to final residue accumulation at 29 Angstroms there isan increase in the numbers hydrophobic residues collected which includesthirty leucine and valine residues.

FIGS. 4A-4D illustrate a distribution of residue centroids andhydophobicity profiles of the subunit C of the ATP synthase fromEscherichia coli, 1 C 17. FIG. 4A illustrates a view of along the C3symmetry axis normal to the plane of the membrane. FIG. 4B illustrates aview canted by 80 degrees from the symmetry axis. FIG. 4C illustratesthe hydrophobicity profile, H(d), calculated with the Eisenberghydrophobicity scale. FIG. 4D illustrates the hydrophobicity profile,H(d), calculated with the GES hydrophobicity scale. The solid lines inFIGS. 4C and 4D are calculated in steps of one Angstroms.

Specifically, FIG. 4A shows the truncated residue centroid distributionof 1 C 17 with a view along the normal to the plane of the membrane. Thetruncated helices of all twelve chains are approximately perpendicularto the plane of the membrane. The slight deviation from cylindricalsymmetry is partially accounted for by the presence of the M chain. Thischain which is adjacent to the helical bundle with approximatecylindrical symmetry, has been deleted in the present calculation.

Since the truncated structure exhibits approximate cylindrical symmetry,profiling was performed with a cylinder of elliptical cross section. Thelargest ellipse shown in FIG. 4A, enclosing all residue centroids, has amajor principal axis of 35 Angstroms. The hydrophobicity profiles, FIGS.4C and 4D, have shapes qualitatively similar to Bacteriorhodopsin. Theprofiles show an initial increase in hydrophobicity after collection ofthe first three residues at 14 Angstroms.

Between the range of values from 17 to 24 Angstroms, there is asignificant reduction in the hydrophobic content of the residuescollected. This is the range of distances between the two inner ellipsesof FIG. 4A, and is the range of distances between the inner and outersets of nested helices. The final region of residue accumulation,between the values of 24 and 35 Angstroms, displays an increase inhydrophobic residue content as the protein-lipid interface from withinthe bilayer is approached.

Of the eleven, 1 C3W and 1 C 17 are the only structures with approximatecylindrical symmetry and with helical axes lying near the normal to thelipid bilayer. Profiles about the cylindrical axes with such orientationwill reflect the demarcation between α-helical nested structuresdifferently from helices that are canted with respect to the bilayersurface. Based on these profiles, it may be surmised that a region ofdecreasing hydrophobic content may be a general feature of the residuedistribution between the nested α-helical bundles of transmembraneproteins.

FIG. 5 is a view of the helices of the truncated mechanosensitive ionchannel, 1MSL, along the normal to the membrane surface. LikeBacteriorhodopsin there are sets of interior and exterior helices, whichare canted, however, with respect to the membrane surface.

FIGS. 6A-6D illustrate a distribution of residue centroids andhydophobicity profiles of the gated mechanosensitive ion channel fromMycobacterium tuberculosis, 1MSL. FIG. 6A illustrates a view of alongthe C3 symmetry axis normal to the plane of the membrane. FIG. 6Billustrates a view canted by 80 degrees from the symmetry aids. FIG. 6Cillustrates the hydrophobicity profile, H(d), calculated with theEisenberg hydrophobicity scale. FIG. 6D illustrates the hydrophobicityprofile, H(d), calculated with the GES hydrophobicity scale. The solidand dashed lines in FIGS. 6C and 6D are calculated in steps of one andtwo Angstroms, respectively.

A view, along the normal to the surface of the bilayer, of thedistribution of residue centroids, FIG. 6A, reveals a region ofseparation between the centroids of the interior and exterior helices.This region, from ten to fifteen Angstroms is between the two innercircles shown in FIG. 6A. FIGS. 6C and 6D show a decrease in residuehydrophobic content with increasing distance within this region, whichis similar to that observed for 1 C3W and 1 C 17.

The prominent increase in hydrophobic content over the range of interiordistances is, however, not observed. With increasing radial distance inthe region of the multimer proximate to the proteinlipid interface, oneobserves increasing residue hydrophobic content. Since the truncatedtransmembrane protein structure of the potassium ion channel, 1 K4D, hasan overall conical shape, a cone has been chosen as the profilinggeometry. The pitch of the cone is chosen visually to register closelywith the exterior distribution of residue centroid locations. As hasbeen mentioned, the nested conical contours will correlate with lipidexposure for residues that are near the protein-lipid boundary.

FIGS. 7A-7D illustrate a distribution of residue centroids andhydophobicity profiles of the potassium ion channel from Streptomyceslividans, 1K4D. FIG. 7A illustrates a view of along the symmetry axisnormal to the plane of the membrane. FIG. 7B illustrates a view cantedby 80 degrees from the symmetry axis. FIG. 7C illustrates thehydrophobicity profile, H (d), calculated with the Eisenberghydrophobicity scale. FIG. 7D illustrates the hydrophobicity profile,H(d), calculated with the GES hydrophobicity scale. The solid and dashedlines in FIGS. 7C and 7D are calculated in steps of one and twoAngstroms, respectively. d is the radius of the smallest radial crosssection of the cone.

The protein has fourfold symmetry about the normal to the membranesurface and the cone is created with varying spherical cross sectionsalong the conical axis. FIG. 7B is a view of the distribution ofcentroids along a direction tilted by 80 degrees from the normal to themembrane surface. The circles of varying radii delineate the cone justlarge enough to enclose all of the residue centroids. FIG. 7A is a viewalong the normal to the membrane surface. The set of circular crosssections shown, differs from previous sets shown since cross sectionsfor only one profiling conical geometry have been displayed. Thedifferent circles delineate the different conical cross-sections of thecone that just enclose all of the centroids.

FIGS. 7C and 7D show the accumulated hydrophobicity, H(d), as a functionof the radius, d. d is the radius of the smallest circular cross sectionof each of the cones of the nested set of conical structures. Thesignificant drop in hydrophobic content between 10 and 11 angstroms isdue to the collection of four Arginine residues. Both residuehydrophobicity scales yield an inverted profile with respect to thatfound for the thirty globular soluble proteins. This again indicatesthat residues nearer to the protein-lipid interface have increasedhydrophobic character with respect to the interior residues.

As the profiling geometry increases in extent over the structureslacking symmetry, it sweeps out spatial regions within nested ellipsoidsthat bear little or no structural resemblance to each other.Consequently, a detailed description of the correspondence betweenprofile features and structural features would be extensive and will notbe provided. Profiles of these structures, FIGS. 8A-D through FIGS.14A-D, are mainly inverted with respect to that found for the solubleproteins.

FIGS. 8A-8D illustrate a distribution of residue centroids andhydophobicity profiles of the photosynthetic reaction center,Rhodobacter sphaeroides, 1PCR. FIG. 8A illustrates a view along the axisnormal to the plane of the membrane. FIG. 8B illustrates a view cantedby 80 degrees from the symmetry axis. FIG. 8C illustrates thehydrophobicity profile, H(d), calculated with the Eisenberghydrophobicity scale. FIG. 8D illustrates the hydrophobicity profile,H(d), calculated with the GES hydrophobicity scale. The solid and dashedlines in FIGS. 8C and 8D are calculated in steps of one and fourAngstroms, respectively.

FIGS. 9A-9D illustrate a distribution of residue centroids andhydophobicity profiles of the Cytochrome-C Oxidase, 1EHK. FIG. 9Aillustrates a view along the axis normal to the plane of the membrane.FIG. 9B illustrates a view canted by 80 degrees from the symmetry axis.FIG. 9C illustrates the hydrophobicity profile, H(d), calculated withthe Eisenberg hydrophobicity scale. FIG. 9D illustrates thehydrophobicity profile, H(d), calculated with the GES hydrophobicityscale. The solid and dashed lines in FIGS. 9C and 9D are calculated insteps of one and two Angstroms, respectively.

FIGS. 10A-10D illustrate a distribution of residue centroids andhydophobicity profiles of the Aqpl Water Channel, 1J4N. FIG. 10Aillustrates a view along the axis normal to the plane of the membrane.FIG. 10B illustrates a view canted by 80 degrees from the symmetry axis.FIG. 10C illustrates the hydrophobicity profile, H(d), calculated withthe Eisenberg hydrophobicity scale. FIG. 10D illustrates thehydrophobicity profile, H(d), calculated with the GES hydrophobicityscale. The solid and dashed lines in FIGS. 10C and 10D are calculated insteps of one and two Angstroms, respectively.

FIGS. 11A-11D illustrate a distribution of residue centroids andhydophobicity profiles of the Bacterial Abc Transporter, 1L7V. FIG. 11Aillustrates a view along the axis normal to the plane of the membrane.FIG. 11B illustrates a view canted by 80 degrees from the symmetry axis.FIG. 11C illustrates the hydrophobicity profile, H (d), calculated withthe Eisenberg hydrophobicity scale. FIG. 11D illustrates thehydrophobicity profile, H(d), calculated with the GES hydrophobicityscale. The solid and dashed lines in FIGS. 11C and 11D are calculated insteps of one and two Angstroms, respectively.

FIGS. 12A-12D illustrate a distribution of residue centroids andhydophobicity profiles of E. Coli Quinol-Fumarate Reductase, 1KF6. FIG.12A illustrates a view along the axis normal to the plane of themembrane. FIG. 12B illustrates a view canted by 80 degrees from thesymmetry axis. FIG. 12C illustrates the hydrophabicity profile, H(d),calculated with the Eisenberg hydrophobicity scale. FIG. 12D illustratesthe hydrophobicity profile, H(d), calculated with the GES hydrophobicityscale. The solid and dashed lines in FIGS. 12C and 12D are calculated insteps of two and three Angstroms, respectively.

FIGS. 13A-13D illustrates a distribution of residue centroids andhydophobicity profiles of the Cytochrome Bc 1 Complex, 1 BE3. FIG. 13Aillustrates a view along the axis normal to the plane of the membrane.FIG. 13B illustrates a view canted by 80 degrees from the symmetry axis.FIG. 13C illustrates the hydrophobicity profile, H(d), calculated withthe Eisenberg hydrophobicity scale. FIG. 13D illustrates thehydrophobicity profile, H(d), calculated with the GES hydrophobicityscale. Both solid and dashed lines in FIGS. 13C and 13D are calculatedin steps of two Angstroms. The solid line is calculated with theinclusion of the D helical chain. The dashed line is calculated withdeletion of this chain.

FIGS. 14A-14D illustrate a distribution of residue centroids andhydophobicity profiles of the Photosynthetic Reaction Center:Photosystem I, 1JB0. FIG. 14A illustrates a view along the axis normalto the plane of the membrane. FIG. 14B illustrates a view canted by 80degrees from the symmetry axis. FIG. 14C illustrates the hydrophobicityprofile, H(d), calculated with the Eisenberg hydrophobicity scale. FIG.14D illustrates the hydrophobicity profile, H(d), calculated with theGES hydrophobicity scale. The solid and dashed lines in FIGS. 14C and14D are calculated in steps of two and five Angstroms, respectively.

It should be noted that of these structures (e.g., FIGS. 8A-D through14A-D), three show little change or a decrease in hydrophobic content atthe farthest distances from the interior. These are the structures, 1K6,1BE3, and 1JB0, FIGS. 12A-D through 14A-D. These particular profilefeatures are not representative of residue accumulation along the majorfraction of the periphery of the protein-lipid interface.

The truncated helical bundle, 1 KF6, in the range of 19 angstroms tocomplete residue collection at 23 angstroms (e.g., see FIG. 12A)collects only few residues at two different spatial locations. Theseresidues have, on average, a hydrophobic content comparable to theaverage hydrophobicity of the entire multimer. The residues collected inthe range of values from 15 to 19 angstroms are greater in number andspan the major fraction of the protein-lipid interface. Over this rangeof distances the hydrophobic residue content, on average, increases asthe interface is approached.

The profiles of the 1BE3 multimer, the solid lines in FIGS. 13C and 13D,show a decrease in hydrophobic content over a range of distances, 26Angstroms to complete residue collection at 32 Angstroms. This behavioris a consequence of the residues of the helical D chain having anaverage value of hydrophobicity that is less than that of the entiremultimer. Profiling the structure with deletion of the D chain yieldsthe dashed profiles shown in FIGS. 13C and 13D. The major fraction ofthe periphery of the multimer then shows increasing hydrophobic contentas the interface with the lipid is approached.

The profiles, FIGS. 14C and 14D, of the multimer, 1JB0, the moststructurally complex structure examined, with multiple chains, show anarrow range of decreasing hydrophobic content with distance close tothe lipid interface. This range of distances is illustrated in FIG. 14A.It is the narrow range of distances between the two closest ellipses.Averages over this range of distances arise from accumulation overseveral different local regions near the interface.

The decrease in hydrophobic content at a distance of 56 Angstroms is dueto the collection of one arginine and one lysine residue in differentlocal regions. Both residues are not only near the protein-lipidinterface within the bilayer but in the vicinity of the bilayer surfaceas well. From 56 Angstroms to final residue collection at 62 Angstromsthere is an increase in hydrophobic content. Differences in the profilesin this narrow spatial region dependent upon windowing size and thechoice of the hydrophobicity scale should also be noted.

The profiles of 1BE3 and 1JB0 emphasize that the distribution of residuehydrophobicity exhibits variations not only in an “inside-outside” orradial direction but in an angular direction as well, near to and alongthe periphery of the protein-lipid interface. Such variations can beinvestigated by profiling along the protein-lipid periphery.

As has been seen, local regions in the vicinity of the protein-lipidinterface may be of lesser hydrophobic content than the multimericaverage. It is of interest that so few such variations have been seen inthe eleven structures examined. Finally, when profiling a complexstructure with multiple helical chains, the baseline for comparison isimportant, e.g., which of the chains are to be chosen to provide thereference value of hydrophobicty against which local variations are tobe compared.

A statistical advantage of collecting the values of residuehydrophobicity within a profiling surface that increases in sizeinvolves a reduction in the fluctuations about the mean, compared withcollecting the values of residue hydrophobicity within each shellbounded by adjacent nested profiling surfaces. The total residuehydrophobicity within each shell divided by the numbers of residues inthe shell, calculated with increasing distance from the axial center ofthe profiling geometry, provides the residue hydrophobicity density,ρ(d), as a function of distance, d, from the center of the structure.

Despite the fluctuations in value, this density is of interest. Thisdensity is illustrated, for example, in FIGS. 15A-F for the foursymmetric α-helical transmembrane bundles, over the larger half of theradial distances, d, from the cylindrical axis; the range of distancesnearer the protein lipid interface. The solid lines are the results ofcalculations for shells of one Angstrom in thickness. The dashed linesare for shells of two Angstroms thickness. It should be noted that onaverage, despite the fluctuations in value, the density of the fourtransmembrane structures exhibits increasing hydrophobic content as theprotein lipid interface is approached.

FIGS. 15E and 15F, pertaining to the soluble proteins, 1AKZ and 3PBG,have been included to highlight the different behavior of theseproteins. For these proteins, the hydrophobic content decreases as theprotein lipid interface is approached. These results obtained for 1AKZand 3PBG are typical of the thirty soluble proteins previously profiled.

As noted from the FIGS. 15A-15F, however, the spatial decrease in thehydrophobic densities of the soluble proteins, 1AKZ and 3PBG, is morepronounced than the increase observed for the transmembrane structures.Comparing peak height amplitudes of the accumulated profiles of thesoluble proteins with the amplitudes of the peak valleys of theα-helical structures generally highlights this more modest segregationof the residue hydrophobic content of the transmembrane bundles.

The terminology, “inside-out”, had been used previously in connectionwith a comparison between the spatial hydrophobicity distributions oftransmembrane and soluble globular proteins. For the Eisenberghydrophobicity scale, which is a consensus set of values thatapproximates the free energy of transfer of the side chain of the aminoacid from water to an a polar environment, the average value per residueof thirty soluble globular proteins is −0.13 kcal/mole.

Further, the average value per residue of the four symmetrictransmembrane bundles is 0.27 kcal/mole. The difference between thesetwo values is comparable to the difference between the values ofThreonine and Alanine on this scale.

The inventor further calculated the differences about the mean value ofhydrophobicity for each of the structures. This requires aredistribution of the individual values of residue hydrophobicity foreach structure, a result achieved by scaling the values of residuehydrophobicity such that the net hydrophobicity of each structurevanished.

FIG. 16 provides Table 1 which lists the shifted and normalized valuesof amino acid hydrophobicity that had provided the values of thedensities, ρ(d). It should be noted that a significant difference existsbetween the values for the soluble and transmembrane structures, as wellas a range of values of opposite sign, within the lines drawn. These arethe values that yield what might be called an “inside-out” distributionof the hydrophobic density of the four symmetric transmembrane bundlesrelative to the soluble proteins, 1AKZ and 3PBG. These values are ameasure of differences about averages, with averages that are verydifferent. The distributions are not “inside-out” in the traditionalsense in which each residue is considered to have a fixed polar or apolar identity.

In short, the present invention may be used to examine the spatialdistribution of transmembrane residue hydrophobicity from a perspectivethat is different from the point of view of conventional calculations.The spatial profile may be obtained directly, without reference tosolvent-lipid exposure. This provides a view of the variation of residuehydrophobicity from the interior to the exterior of the α-helicalbundles buried within the surrounding lipid.

Further, the scaling the residue hydrophobicity for each multimerenables variations about the mean value of hydrophobicity over thespatial extent of the structure to be simply identified. This alsoenables a comparison of the profiles over the spatial extent ofdifferent structures with average values of hydrophobicity that aredifferent. Such a procedure had previously identified comparable lengthscale features of the profiles of thirty soluble globular proteins ofarbitrary structure and size.

The profiles of the α-helical buried bundles, while exhibiting certaindifferences, exhibit a comparable length scale feature as well. This isthe onset of the decrease in hydrophobic residue content at distancesfrom the interior that are at roughly half the spatial extent of thebundle. Consequently, the profiles are mainly inverted with respect tothe profiles of the soluble globular proteins. The region proximate tothe protein-lipid interface, that had generated previous contention,generally exhibits the increase in average residue hydrophobic contentidentified by previous calculations. The profiling of the structureslacking symmetry show that such increase need not occur in every localregion proximate to the protein-lipid periphery.

Referring again to the drawings, FIG. 17 illustrates a method 1700 ofspatially profiling a hydrophobicity distribution for a transmembraneprotein. The method 1700 includes scaling hydrophobicity values (1710)for the transmembrane protein to generate scaled hydrophobicity values,and spatially profiling (1720) a hydrophobicity distribution for thetransmembrane protein based on the scaled hydrophobicity values.Specifically, the method 1700 may be performed in a manner which issimilar to that outlined above with respect to the system 100.

Referring now to FIG. 18, system 1800 illustrates a typical hardwareconfiguration which may be used to implement the inventive system andmethod for spatially profiling a hydrophobicity distribution for atransmembrane protein. The configuration has preferably at least oneprocessor or central processing unit (CPU) 1811. The CPUs 1811 areinterconnected via a system bus 1812 to a random access memory (RAM)1814, read-only memory (ROM) 1816, input/output (I/O) adapter 1818 (forconnecting peripheral devices such as disk units 1821 and tape drives1840 to the bus 1812), user interface adapter 1822 (for connecting akeyboard 1824, mouse 1826, speaker 1828, microphone 1832, and/or otheruser interface device to the bus 1812), a communication adapter 1834 forconnecting an information handling system to a data processing network,the Internet, and Intranet, a personal area network (PAN), etc., and adisplay adapter 1836 for connecting the bus 1812 to a display device1838 and/or printer 1839. Further, an automated reader/scanner 1841 maybe included. Such readers/scanners are commercially available from manysources.

In addition to the system described above, a different exemplary aspectof the invention includes a computer-implemented method for performingthe above method. As an example, this method may be implemented in theparticular environment discussed above.

Such a method may be implemented, for example, by operating a computer,as embodied by a digital data processing apparatus, to execute asequence of machine-readable instructions. These instructions may residein various types of signal-bearing media.

Thus, this exemplary aspect of the present invention is directed to aprogrammed product, including signal-bearing media tangibly embodying aprogram of machine-readable instructions executable by a digital dataprocessor to perform the above method.

Such a method may be implemented, for example, by operating the CPU 1811to execute a sequence of machine-readable instructions. Theseinstructions may reside in various types of signal bearing media.

Thus, this exemplary aspect of the present invention is directed to aprogrammed product, comprising signal-bearing media tangibly embodying aprogram of machine-readable instructions executable by a digital dataprocessor incorporating the CPU 1811 and hardware above, to perform themethod of the invention.

This signal-bearing media may include, for example, a RAM containedwithin the CPU 1811, as represented by the fast-access storage forexample. Alternatively, the instructions may be contained in anothersignal-bearing media, such as a magnetic data storage diskette 1900(FIG. 19), directly or indirectly accessible by the CPU 1811.

Whether contained in the computer server/CPU 1811, or elsewhere, theinstructions may be stored on a variety of machine-readable data storagemedia, such as DASD storage (e.g, a conventional “hard drive” or a RAIDarray), magnetic tape, electronic read-only memory (e.g., ROM, EPROM, orEEPROM), an optical storage device (e.g., CD-ROM, WORM, DVD, digitaloptical tape, etc.), paper “punch”cards, or other suitablesignal-bearing media including transmission media such as digital andanalog and communication links and wireless. In an illustrativeembodiment of the invention, the machine-readable instructions maycomprise software object code, complied from a language such as C+, C++etc.

With its unique and novel features, the present invention provides asystem and method which enables a determination of the spatialdistribution of hydrophobicity over an entire multimeric extent, notonly in the region proximate to the protein lipid interface.

While the invention has been described in terms of one or more exemplaryembodiments, those skilled in the art will recognize that the inventioncan be practiced with modification within the spirit and scope of theappended claims. Specifically, one of ordinary skill in the art willunderstand that the drawings herein are meant to be illustrative, andthe design of the inventive assembly is not limited to that disclosedherein but may be modified within the spirit and scope of the presentinvention.

Further, Applicant's intent is to encompass the equivalents of all claimelements, and no amendment to any claim the present application shouldbe construed as a disclaimer of any interest in or right to anequivalent of any element or feature of the amended claim.

1. A system for spatially profiling a hydrophobicity distribution for atransmembrane protein, said system comprising: a scaler which generatesscaled hydrophobicity values for said transmembrane protein; a profilerwhich spatially profiles a hydrophobicity distribution for saidtransmembrane protein based on said scaled hydrophobicity value; and adisplay device for displaying output of said profiler, wherein saidprofiler generates a hydrophobicity distribution profile for saidprotein using a predetermined profiling geometry which is selected toapproximate an external shape of said protein within a lipid bilayer,and wherein said profiler comprises: an identifier for identifying aresidue external to said membrane and removing said residue to obtain atruncated structure comprising plural residue side-chains which areinternal to said membrane; and a calculator which: calculates pluralresidue centroids for said plural residue side-chains; calculates adistribution of said plural residue centroids using a series of nestedshapes which are consistent with said profiling geometry and providecontours about an axis that correlate with a lipid exposure for theresidues proximate to a boundary between said protein and said lipidbilayer; and obtains a geometric center for said distribution of saidplural residue centroids, said hydrophobicity profile being about saidaxis through said geometric center and normal to a plane of saidmembrane.
 2. The system according to claim 1, wherein said scaledhydrophobicity values comprise shifted and scaled hydrophobicity valuesof residue hydrophobicity.
 3. The system according to claim 1, whereinsaid profiler spatially profiles said hydrophobicity distribution over aspatial extent of said transmembrane protein.
 4. The system according toclaim 1, wherein said scaled hydrophobicity values are derived from datapertaining to a structure of said transmembrane protein.
 5. The systemaccording to claim 1, wherein said identifier identifies a residueexternal to said membrane, using at least one of hydrophobicity slidingwindow analysis and visual inspection.
 6. The system according to claim5, wherein said profiler further comprises a residue remover whichremoves said residue to obtain said truncated structure.
 7. The systemaccording to claim 1, wherein a structure of said transmembrane proteinis asymmetrical about said axis, and said predetermined profilinggeometry comprises an elliptical cylinder.
 8. The system according toclaim 7, wherein said scaler scales said hydrophobicity values to obtainsaid scaled hydrophobicity values, using a scale that segregates aminoacid values of hydrophobicity into a polar, polar uncharged and polarcharged residues.
 9. The system according to claim 8, wherein saidscaler shifts said scaled hydrophobicity values to provide shifted andscaled values having a value of zero hydrophobicity when a predeterminednumber of residues of each truncated structure are collected, saidshifted and scaled values being scaled to provide a standard deviationof unity for each truncated structure.
 10. The system according to claim9, wherein said profiler accumulates said shifted and scaled values ofresidue hydrophobicity as a function of an increasing size of eachnested shape of a profiling geometry until a largest shape encapsulatesa predetermined number of the residues, to generate an accumulatedspatial distribution of residue hydrophobicity given by a function H(d)which is a sum of residue hydophobicity values within the circular,elliptical or conical cylinder of radius d.
 11. The system according toclaim 10, wherein said profiler obtains a hydrophobicity profile bycalculating the values of H(d).
 12. The system according to claim 11,wherein if H(d) increases, the average hydrophobic value of the residuescollected over a shell of width one or more Angstroms is greater thanthe average value of residue hydrophobicity for the entire structure,and wherein if H(d) decreases, the average hydrophobic value of theresidues collected over a shell of width one or more Angstroms is lessthan the average value of residue hydrophobicity for the entirestructure.
 13. The system according to claim 10, wherein saidaccumulated spatial distribution comprises a set of sequential values ofaccumulated residue hydrophobicity with increasing distance from acenter of the structure to the protein/lipid interface within a bilayer,said set of sequential values comprising a zero-order moment profile ofthe residue hydrophobicity from the interior to the exterior of thestructure.
 14. The system according to claim 1, further comprising: adatabase which stores data pertaining to a structure of saidtransmembrane protein; and an input device for inputting andmanipulating said data.
 15. A method of spatially profiling ahydrophobicity distribution for a transmembrane protein, said methodcomprising: scaling hydrophobicity values for said transmembrane proteinto generate scaled hydrophobicity values; spatially profiling ahydrophobicity distribution for said transmembrane protein based on saidscaled hydrophobicity values; and a display device for displaying outputof said profiler, wherein said spatially profiling comprises generatinga hydrophobicity distribution profile for said protein using apredetermined profiling geometry which is selected to approximate anexternal shape of said protein within a lipid bilayer, said generatingsaid hydrophobicity distribution profile comprising: identifying aresidue external to said membrane and removing said residue to obtain atruncated structure comprising plural residue side-chains which areinternal to said membrane; calculating plural residue centroids for saidplural residue side-chains; calculating a distribution of said pluralresidue centroids using a series of nested shapes which are consistentwith said profiling geometry and provide contours about an axis thatcorrelate with a lipid exposure for the residues proximate to a boundarybetween said protein and said lipid bilayer; and obtaining a geometriccenter for said distribution of said plural residue centroids, saidhydrophobicity profile being about said axis through said geometriccenter and normal to a plane of said membrane.
 16. The method accordingto claim 15, wherein said identifying a residue external to saidmembrane, comprises using hydrophobicity sliding window analysis andvisual inspection.
 17. The method according to claim 15, wherein saidscaling said hydrophobicity values comprises scaling hydrophobicityvalues to obtain said scaled hydrophobicity values, using a scale thatsimilarly segregates amino acid values of hydrophobicity into a polar,polar uncharged and polar charged residues.
 18. The method according toclaim 17, wherein said scaling said hydrophobicity values furthercomprises shifting said scaled hydrophobicity values to provide shiftedand scaled values having a value of zero hydrophobicity when apredetermined number of residues of each truncated structure arecollected, said shifted and scaled values being scaled to provide astandard deviation of unity for each truncated structure.
 19. The methodaccording to claim 18, wherein said spatially profiling furthercomprises accumulating said shifted and scaled values of residuehydrophobicity as a function of increasing size of each nested shape ofthe profiling geometry until a largest shape encapsulates apredetermined number of the residues, to generate an accumulated spatialdistribution of residue hydrophobicity given by the function H(d) whichis the sum of the values of residue hydophobicity within the circular,elliptical or conical cylinder of radius d.
 20. The method according toclaim 19, wherein said spatially profiling further comprises obtaining ahydrophobicity profile by calculating the values of H(d).
 21. The systemaccording to claim 1, wherein said scaled hydrophobicity values compriseinformation regarding a proposed three-dimensional transmembranestructure of said transmembrane protein.
 22. The system according toclaim 21, wherein said profiler profiles said hydrophobicitydistribution for said transmembrane protein to validate said proposedthree-dimensional transmembrane structure.